Monoclonal antibody produced by heterologous indirect enzyme-linked immunosorbent assay and its application for parathion residue determination in agricultural and environmental samples

Abstract

A heterologous indirect enzyme-linked immunosorbent assay (ELISA) was developed, which was based on monoclonal antibody (Mab) to determine parathion residue in agricultural and environmental samples. Eight Mabs were produced by introducing the heterologous indirect ELISA into the screening procedure. It was shown that these Mabs had more broad-reactivity with twenty competitors than that of 5H7 (Mab produced by homologous screening). So it became much easier using these new Mabs to develop heterologous immunoassays. In addition, all the developed heterologous ELISAs could be used to determine parathion residue in semiquantitative or quantitative level, and their detection limitation (LOD) was around 2 ng/mL. Compared with the previous heterologous ELISA (hapten 11/5H7) with IC₅₀ of 13.3 ng/mL, a better heterologous ELISA (hapten 11/1E1) was obtained with IC₅₀ of 3.8 ng/mL, which improved the sensitivity 3.5 times. Finally, the latter was applied to parathion residue determination in spiked agricultural and environmental samples without extraction or cleanup. The average recoveries of parathion added to water, soil, cucumber, Chinese cabbage and carrot were between 80.4 % and 111.8 %. The LOD for water and soil samples was 5 ng/mL, and the LOD for cucumber, rice and corn samples was 10 ng/mL.

Keywords:

• Monoclonal antibody
• heterologous immunoassay
• enzyme-linked immunosorbent assay
• parathion

Acknowledgments

This work was supported by the National Natural Science Foundation of China (NSFC No. 30671391) and the Key Projects in the National Science & Technology Pillar Program during the Eleventh Five-year Plan Period (NO2009BADB9B09).

Notes

^a •••: conjugates showed high reactivity (A ≥ 1.5) with the antibody; ••○: conjugates showed middle reactivity (1.0 ≤ A < 1.5) with the antibody; •○○: conjugates showed low reactivity (0.5 ≤ A < 1.0) with the antibody; ○○○: conjugates showed no reactivity (A < 0.5) with the antibody.

^bMab 5H7 was produced in our previous work.

^cBroad-reactivity, the number of the designed competitors which could be recognized by the Mab.

^a: Results were determined by heteroglogous ELISA (hapten 11/1E1);

^b: Results were determined by GC.